PROJECT SUMMARY The widespread misuse of prescription -opioid receptor (MOPR) opioids, opioid addiction, and overdose has underscored the need to develop effective, non-addicting medications to treat pain. Chronic pain is a major factor contributing to insomnia, and sleep disruption due to chronic pain causes patients to seek relief, exacerbating the drive for prescription opioids. In Opioid Use Disorder (OUD), withdrawal from opiates induces insomnia, posing an additional challenge for successful abstinence. The central hypothesis of this proposal is that treatment of opioid withdrawal-induced insomnia with nociceptin/orphanin FQ receptor (NOPR) agonists will mitigate the drive for opiate use. Since NOPR agonists have shown promise as modulators of the antinociceptive and rewarding effects of opiates, the N/OFQ?NOPR system may be a pathway through which to interrupt the pain => opiate use => withdrawal => insomnia => opiate use cycle. We have found that NOPR agonists potently induce sleep and increase EEG delta power in rodents. Consequently, NOPR agonists may have a dual role to reduce opiate use through their analgesic activity and by enhancing sleep. A major component of the arousal and withdrawal circuitries resides in the locus coeruleus (LC), a noradrenergic center that expresses MOPRs, initiates arousal, and mediates many opioid withdrawal phenotypes. We will determine whether and how the N/OFQ-NOPR system engages LC circuits to reduce arousal and insomnia- related phenotypes and assess the hypotheses that (1) the N/OFQ-NOPR system is a component of the endogenous sleep/wake regulatory system and (2) NOPR agonists can act as therapeutic interventions to reduce opiate use. To determine whether NOPR activation mitigates morphine withdrawal-induced insomnia in Specific Aim 1, we will characterize sleep/wake disruption in a mouse model of morphine withdrawal, evaluate the efficacy of NOPR agonism and genetic disruption of NOPR on withdrawal-induced insomnia, determine whether NOPR agonists are effective hypnotics in primates as well as rodents, and evaluate the role of the N/OFQ-NOPR system in sleep/wake control. In Aim 2, using cutting-edge circuit dissection tools and newly- developed transgenic models that allow for specific manipulation of the N/OFQ system in vivo, we will determine NOPR localization within identified LC cell groups and LC inputs known to regulate sleep/wake, measure the activity of LC-NE and LC-GABA neurons during sleep/wake using fiber photometry, use Pnoc- ires-cre mice to locally manipulate these specific N/OFQ-NOPR circuits, and utilize viral CRISPR/Cas9-based tools and conditional PNOC mice to selectively remove the endogenous N/OFQ peptide from the LC in specific cell types and determine the consequences on sleep-wake regulation. Together, these approaches will help determine whether the manipulation of N/OFQ-NOPR system is a potentially novel pathway to treat morphine withdrawal-induced insomnia and help break the cycle of opiate dependence.